scholarly journals Membrane-anchored growth factor, HB-EGF, on the cell surface targeted to the inner nuclear membrane

2008 ◽  
Vol 180 (4) ◽  
pp. 763-769 ◽  
Author(s):  
Miki Hieda ◽  
Mayumi Isokane ◽  
Michiko Koizumi ◽  
Chiduru Higashi ◽  
Taro Tachibana ◽  
...  
Keyword(s):  
Growth Factor ◽  
Cell Surface ◽  
Nuclear Envelope ◽  
Nuclear Membrane ◽  
Transmembrane Protein ◽  
Active Form ◽  
Type I ◽  
Transcriptional Repressors ◽  
Sequence Element ◽  
Inner Nuclear Membrane

Heparin-binding EGF-like growth factor (HB-EGF) is synthesized as a type I transmembrane protein (proHB-EGF) and expressed on the cell surface. The ectodomain shedding of proHB-EGF at the extracellular region on the plasma membrane yields a soluble EGF receptor ligand and a transmembrane-cytoplasmic fragment (HB-EGF-CTF). The cytoplasmic domain of proHB-EGF (HB-EGF-cyto) interacts with transcriptional repressors to reverse their repressive activities. However, how HB-EGF-cyto accesses transcriptional repressors is yet unknown. The present study demonstrates that, after exposure to shedding stimuli, both HB-EGF-CTF and unshed proHB-EGF translocate to the nuclear envelope. Immunoelectron microscopy and digitonin-permeabilized cells showed that HB-EGF-cyto signals are at the inner nuclear membrane. A short sequence element within the HB-EGF-cyto allows a transmembrane protein to localize to the nuclear envelope. The dominant-active form of Rab5 and Rab11 suppressed nuclear envelope targeting. Collectively, these data demonstrate that membrane-anchored HB-EGF is targeted to the inner nuclear membrane via a retrograde membrane trafficking pathway.

scholarly journals Opposing roles for distinct LINC complexes in regulation of the small GTPase RhoA

2017 ◽  
Vol 28 (1) ◽  
pp. 182-191 ◽  
Author(s):  
Ketan Thakar ◽  
Christopher K. May ◽  
Anna Rogers ◽  
Christopher W. Carroll
Keyword(s):  
Membrane Protein ◽  
Actin Cytoskeleton ◽  
Nuclear Envelope ◽  
Focal Adhesion ◽  
Nuclear Membrane ◽  
Active Form ◽  
Small Gtpase ◽  
Inner Nuclear Membrane ◽  
Inner Nuclear Membrane Protein ◽  
Nuclear Membrane Protein

Linker of Nucleoskeleton and Cytoskeleton (LINC) complexes span the nuclear envelope and transduce force from dynamic cytoskeletal networks to the nuclear lamina. Here we show that LINC complexes also signal from the nuclear envelope to critical regulators of the actin cytoskeleton. Specifically, we find that LINC complexes that contain the inner nuclear membrane protein Sun2 promote focal adhesion assembly by activating the small GTPase RhoA. A key effector in this process is the transcription factor/coactivator complex composed of SRF/Mkl1. A constitutively active form of SRF/Mkl1 was not sufficient to induce focal adhesion assembly in cells lacking Sun2, however, suggesting that LINC complexes support RhoA activity through a transcription-independent mechanism. Strikingly, we also find that the inner nuclear membrane protein Sun1 antagonizes Sun2 LINC complexes and inhibits RhoA activation and focal adhesion assembly. Thus different LINC complexes have opposing roles in the transcription-independent control of the actin cytoskeleton through the small GTPase RhoA.

scholarly journals Myne-1, a spectrin repeat transmembrane protein of the myocyte inner nuclear membrane, interacts with lamin A/C

2002 ◽  
Vol 115 (1) ◽  
pp. 61-70 ◽  
Author(s):  
John M. K. Mislow ◽  
Marian S. Kim ◽  
Dawn Belt Davis ◽  
Elizabeth M. McNally
Keyword(s):  
Muscular Dystrophy ◽  
Nuclear Envelope ◽  
Nuclear Membrane ◽  
Transmembrane Domain ◽  
Transmembrane Protein ◽  
Lamin A ◽  
Spectrin Repeat ◽  
Inner Nuclear Membrane ◽  
C Terminus

Mutations in the genes encoding the inner nuclear membrane proteins lamin A/C and emerin produce cardiomyopathy and muscular dystrophy in humans and mice. The mechanism by which these broadly expressed gene products result in tissue-specific dysfunction is not known. We have identified a protein of the inner nuclear membrane that is highly expressed in striated and smooth muscle. This protein, myne-1 (myocyte nuclear envelope), is predicted to have seven spectrin repeats, an interrupted LEM domain and a single transmembrane domain at its C-terminus. We found that myne-1 is expressed upon early muscle differentiation in multiple intranuclear foci concomitant with lamin A/C expression. In mature muscle, myne-1 and lamin A/C are perfectly colocalized, although colocalization with emerin is only partial. Moreover, we show that myne-1 and lamin A/C coimmunoprecipitate from differentiated muscle in vitro. The muscle-specific inner nuclear envelope expression of myne-1, along with its interaction with lamin A/C, indicates that this gene is a potential mediator of cardiomyopathy and muscular dystrophy.

scholarly journals Unrestrained ESCRT-III drives chromosome fragmentation and micronuclear catastrophe

2019 ◽  
Author(s):  
Marina Vietri ◽  
Sebastian W. Schultz ◽  
Aurélie Bellanger ◽  
Carl M. Jones ◽  
Camilla Raiborg ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Membrane ◽  
Genome Stability ◽  
Membrane Deformation ◽  
Chromosome Fragmentation ◽  
Inner Nuclear Membrane ◽  
Membrane Rupture ◽  
Membrane Fission ◽  
Stable Association ◽  
Inner Nuclear Membrane Protein

AbstractThe ESCRT-III membrane fission machinery1,2 restores nuclear envelope integrity during mitotic exit3,4 and interphase5,6. Whereas primary nuclei resealing takes minutes, micronuclear envelope ruptures appear irreversible and result in catastrophic collapse associated with chromosome fragmentation and rearrangements (chromothripsis), thought to be a major driving force in cancer development7-10. Despite its importance11-13, the mechanistic underpinnings of nuclear envelope sealing in primary nuclei and the defects observed in micronuclei remain largely unknown. Here we show that CHMP7, the nucleator of ESCRT-III filaments at the nuclear envelope3,14, and the inner nuclear membrane protein LEMD215 act as a compartmentalization sensor detecting the loss of nuclear integrity. In cells with intact nuclear envelope, CHMP7 is actively excluded from the nucleus to preclude its binding to LEMD2. Nuclear influx of CHMP7 results in stable association with LEMD2 at the inner nuclear membrane that licenses local polymerization of ESCRT-III. Tight control of nuclear CHMP7 levels is critical, as induction of nuclear CHMP7 mutants is sufficient to induce unrestrained growth of ESCRT-III foci at the nuclear envelope, causing dramatic membrane deformation, local DNA torsional stress, single-stranded DNA formation and fragmentation of the underlying chromosomes. At micronuclei, membrane rupture is not associated with repair despite timely recruitment of ESCRT-III. Instead, micronuclei inherently lack the capacity to restrict accumulation of CHMP7 and LEMD2. This drives unrestrained ESCRT-III recruitment, membrane deformation and DNA defects that strikingly resemble those at primary nuclei upon induction of nuclear CHMP7 mutants. Preventing ESCRT-III recruitment suppresses membrane deformation and DNA damage, without restoring nucleocytoplasmic compartmentalization. We propose that the ESCRT-III nuclear integrity surveillance machinery is a double-edged sword, as its exquisite sensitivity ensures rapid repair at primary nuclei while causing unrestrained polymerization at micronuclei, with catastrophic consequences for genome stability16-18.

scholarly journals Characterization of Endogenous SERINC5 Protein as Anti-HIV-1 Factor

2019 ◽  
Vol 93 (24) ◽  
Author(s):  
Vânia Passos ◽  
Thomas Zillinger ◽  
Nicoletta Casartelli ◽  
Amelie S. Wachs ◽  
Shuting Xu ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Surface ◽  
Subcellular Localization ◽  
Transmembrane Protein ◽  
Type I ◽  
Accessory Gene ◽  
Protein Levels ◽  
Specificity And Sensitivity ◽  
Hiv 1

ABSTRACT When expressed in virus-producing cells, the cellular multipass transmembrane protein SERINC5 reduces the infectivity of HIV-1 particles and is counteracted by HIV-1 Nef. Due to the unavailability of an antibody of sufficient specificity and sensitivity, investigation of SERINC5 protein expression and subcellular localization has been limited to heterologously expressed SERINC5. We generated, via CRISPR/Cas9-assisted gene editing, Jurkat T-cell clones expressing endogenous SERINC5 bearing an extracellularly exposed hemagglutinin (HA) epitope [Jurkat SERINC5(iHA knock-in) T cells]. This modification enabled quantification of endogenous SERINC5 protein levels and demonstrated a predominant localization in lipid rafts. Interferon alpha (IFN-α) treatment enhanced cell surface levels of SERINC5 in a ruxolitinib-sensitive manner in the absence of modulation of mRNA and protein quantities. Parental and SERINC5(iHA knock-in) T cells shared the ability to produce infectious wild-type HIV-1 but not an HIV-1 Δnef mutant. SERINC5-imposed reduction of infectivity involved a modest reduction of virus fusogenicity. An association of endogenous SERINC5 protein with HIV-1 Δnef virions was consistently detectable as a 35-kDa species, as opposed to heterologous SERINC5, which presented as a 51-kDa species. Nef-mediated functional counteraction did not correlate with virion exclusion of SERINC5, arguing for the existence of additional counteractive mechanisms of Nef that act on virus-associated SERINC5. In HIV-1-infected cells, Nef triggered the internalization of SERINC5 in the absence of detectable changes of steady-state protein levels. These findings establish new properties of endogenous SERINC5 expression and subcellular localization, challenge existing concepts of HIV-1 Nef-mediated antagonism of SERINC5, and uncover an unprecedented role of IFN-α in modulating SERINC5 through accumulation at the cell surface. IMPORTANCE SERINC5 is the long-searched-for antiviral factor that is counteracted by the HIV-1 accessory gene product Nef. Here, we engineered, via CRISPR/Cas9 technology, T-cell lines that express endogenous SERINC5 alleles tagged with a knocked-in HA epitope. This genetic modification enabled us to study basic properties of endogenous SERINC5 and to verify proposed mechanisms of HIV-1 Nef-mediated counteraction of SERINC5. Using this unique resource, we identified the susceptibility of endogenous SERINC5 protein to posttranslational modulation by type I IFNs and suggest uncoupling of Nef-mediated functional antagonism from SERINC5 exclusion from virions.

Function and assembly of nuclear pore complex proteins

1999 ◽  
Vol 77 (4) ◽  
pp. 321-329 ◽  
Author(s):  
Khaldon Bodoor ◽  
Sarah Shaikh ◽  
Paul Enarson ◽  
Sharmin Chowdhury ◽  
Davide Salina ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Nuclear Envelope ◽  
Nuclear Pore Complex ◽  
Nuclear Membrane ◽  
Current View ◽  
Nuclear Pore ◽  
Dynamic Component ◽  
Inner Nuclear Membrane ◽  
Inner Nuclear Membrane Protein ◽  
Nuclear Membrane Protein

Nuclear pore complexes (NPCs) are extremely elaborate structures that mediate the bidirectional movement of macromolecules between the nucleus and cytoplasm. The current view of NPC organization features a massive symmetrical framework that is embedded in the double membranes of the nuclear envelope. It embraces a central channel of as yet ill-defined structure but which may accommodate particles with diameters up to 26 nm provided that they bear specific import/export signals. Attached to both faces of the central framework are peripheral structures, short cytoplasmic filaments, and a nuclear basket assembly, which interact with molecules transiting the NPC. The mechanisms of assembly and the nature of NPC structural intermediates are still poorly understood. However, mutagenesis and expression studies have revealed discrete sequences within certain NPC proteins that are necessary and sufficient for their appropriate targeting. In addition, some details are emerging from observations on cells undergoing mitosis where the nuclear envelope is disassembled and its components, including NPC subunits, are dispersed throughout the mitotic cytoplasm. At the end of mitosis, all of these components are reutilized to form nuclear envelopes in the two daughter cells. To date, it has been possible to define a time course of postmitotic assembly for a group of NPC components (CAN/Nup214, Nup153, POM121, p62 and Tpr) relative to the integral inner nuclear membrane protein LAP2 and the NPC membrane glycoprotein gp210. Nup153, a dynamic component of the nuclear basket, associates with chromatin towards the end of anaphase coincident with, although independent of, the inner nuclear membrane protein, LAP2. Assembly of the remaining proteins follows that of the nuclear membranes and occurs in the sequence POM121, p62, CAN/Nup214 and gp210/Tpr. Since p62 remains as a complex with three other NPC proteins (p58, p54, p45) during mitosis, and CAN/Nup214 maintains a similar interaction with its partner, Nup84, the relative timing of assembly of these additional four proteins may also be inferred. These observations suggest that there is a sequential association of NPC proteins with chromosomes during nuclear envelope reformation and the recruitment of at least eight of these precedes that of gp210. These findings support a model in which it is POM121 rather than gp210 that defines initial membrane-associated NPC assembly intermediates and which may therefore represent an essential component of the central framework of the NPC. Key words: nuclear pore complex, nucleoporin, mitosis, nuclear transport

scholarly journals Tetraspanins TSP-12 and TSP-14 function redundantly to regulate the trafficking of the type II BMP receptor in Caenorhabditis elegans

2020 ◽  
Vol 117 (6) ◽  
pp. 2968-2977
Author(s):  
Zhiyu Liu ◽  
Herong Shi ◽  
Anthony K. Nzessi ◽  
Anne Norris ◽  
Barth D. Grant ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Cell Surface ◽  
Molecular Mechanisms ◽  
Transmembrane Protein ◽  
Expression Patterns ◽  
Bmp Signaling ◽  
Type I ◽  
Type Ii ◽  
Bmp Receptors

Tetraspanins are a unique family of 4-pass transmembrane proteins that play important roles in a variety of cell biological processes. We have previously shown that 2 paralogous tetraspanins in Caenorhabditis elegans, TSP-12 and TSP-14, function redundantly to promote bone morphogenetic protein (BMP) signaling. The underlying molecular mechanisms, however, are not fully understood. In this study, we examined the expression and subcellular localization patterns of endogenously tagged TSP-12 and TSP-14 proteins. We found that TSP-12 and TSP-14 share overlapping expression patterns in multiple cell types, and that both proteins are localized on the cell surface and in various types of endosomes, including early, late, and recycling endosomes. Animals lacking both TSP-12 and TSP-14 exhibit reduced cell-surface levels of the BMP type II receptor DAF-4/BMPRII, along with impaired endosome morphology and mislocalization of DAF-4/BMPRII to late endosomes and lysosomes. These findings indicate that TSP-12 and TSP-14 are required for the recycling of DAF-4/BMPRII. Together with previous findings that the type I receptor SMA-6 is recycled via the retromer complex, our work demonstrates the involvement of distinct recycling pathways for the type I and type II BMP receptors and highlights the importance of tetraspanin-mediated intracellular trafficking in the regulation of BMP signaling in vivo. As TSP-12 and TSP-14 are conserved in mammals, our findings suggest that the mammalian TSP-12 and TSP-14 homologs may also function in regulating transmembrane protein recycling and BMP signaling.

scholarly journals A Rat Pituitary Tumor Cell Line (GH) Expresses Type I and Type II Receptors and Other Cell Surface Binding Protein(s) for Transforming Growth Factor-

1995 ◽  
Vol 270 (2) ◽  
pp. 770-774 ◽  
Author(s):  
Hidetoshi Yamashita ◽  
Toshihide Okadome ◽  
Petra Franzén ◽  
Peter ten Dijke ◽  
Carl-Henrik Heldin ◽  
...  
Keyword(s):  
Growth Factor ◽  
Cell Surface ◽  
Transforming Growth Factor ◽  
Tumor Cell Line ◽  
Protein S ◽  
Type I ◽  
Type Ii ◽  
Surface Binding ◽  
Cell Surface Binding ◽  
Rat Pituitary

scholarly journals The Integral Inner Nuclear Membrane Protein MAN1 Physically Interacts with the R-Smad Proteins to Repress Signaling by the Transforming Growth Factor-β Superfamily of Cytokines

2005 ◽  
Vol 280 (16) ◽  
pp. 15992-16001 ◽  
Author(s):  
Deng Pan ◽  
Luis D. Estévez-Salmerón ◽  
Shannon L. Stroschein ◽  
Xueliang Zhu ◽  
Jun He ◽  
...  
Keyword(s):  
Growth Factor ◽  
Membrane Protein ◽  
Nuclear Membrane ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Activin Signaling ◽  
Inner Nuclear Membrane ◽  
Smad Proteins ◽  
Inner Nuclear Membrane Protein ◽  
Nuclear Membrane Protein

Smad proteins are critical intracellular mediators of the transforming growth factor-β, bone morphogenic proteins (BMPs), and activin signaling. Upon ligand binding, the receptor-associated R-Smads are phosphorylated by the active type I receptor serine/threonine kinases. The phosphorylated R-Smads then form heteromeric complexes with Smad4, translocate into the nucleus, and interact with various transcription factors to regulate the expression of downstream genes. Interaction of Smad proteins with cellular partners in the cytoplasm and nucleus is a critical mechanism by which the activities and expression of the Smad proteins are modulated. Here we report a novel step of regulation of the R-Smad function at the inner nuclear membrane through a physical interaction between the integral inner nuclear membrane protein MAN1 and R-Smads. MAN1, through the RNA recognition motif, associates with R-Smads but not Smad4 at the inner nuclear membrane in a ligand-independent manner. Overexpression of MAN1 results in inhibition of R-Smad phosphorylation, heterodimerization with Smad4 and nuclear translocation, and repression of transcriptional activation of the TGFβ, BMP2, and activin-responsive promoters. This repression of TGFβ, BMP2, and activin signaling is dependent on the MAN1-Smad interaction because a point mutation that disrupts this interaction abolishes the transcriptional repression by MAN1. Thus, MAN1 represents a new class of R-Smad regulators and defines a previously unrecognized regulatory step at the nuclear periphery.

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